SUMMARY While VEGF-targeted therapies are showing promise, new angiogenesis targets are needed to make additional gains. Here, we show that increased Zeste homologue 2 (EZH2) expression in either tumor cells or in tumor vasculature is predictive of poor clinical outcome. The increase in endothelial EZH2 is a direct result of VEGF stimulation by a paracrine circuit that promotes angiogenesis by methylating and silencing vasohibin1 (VASH1). EZH2 silencing in the tumor-associated endothelial cells inhibited angiogenesis mediated by reactivation of VASH1, and reduced ovarian cancer growth, which is further enhanced in combination with EZH2 silencing in tumor cells. Collectively, these data support the potential for targeting EZH2 as an important therapeutic approach. SIGNIFICANCE In this work, we identify EZH2 as a key regulator of tumor angiogenesis. The increase in endothelial EZH2 is a direct result of VEGF stimulation and indicates the presence of a paracrine circuit that promotes angiogenesis. EZH2 silencing in the tumor-associated endothelial cells using siRNA, packaged in the chitosan delivery system, resulted in significant growth inhibition in an orthotopic ovarian cancer model. EZH2 silencing in tumor endothelial cells resulted in decreased angiogenesis that was mediated by increased levels of the angiogenesis inhibitor, vasohibin1 (VASH1). Combined, these data provide a significant conceptual advance in our understanding of the regulation of angiogenesis in ovarian carcinoma and support the potential for targeting EZH2 as a therapeutic approach.

The miR-200 family is well known to inhibit the epithelial–mesenchymal transition, suggesting it may therapeutically inhibit metastatic biology. However, conflicting reports regarding the role of miR-200 in suppressing or promoting metastasis in different cancer types have left unanswered questions. Here we demonstrate a difference in clinical outcome based on miR-200's role in blocking tumour angiogenesis. We demonstrate that miR-200 inhibits angiogenesis through direct and indirect mechanisms by targeting interleukin-8 and CXCL1 secreted by the tumour endothelial and cancer cells. Using several experimental models, we demonstrate the therapeutic potential of miR-200 delivery in ovarian, lung, renal and basal-like breast cancers by inhibiting angiogenesis. Delivery of miR-200 members into the tumour endothelium resulted in marked reductions in metastasis and angiogenesis, and induced vascular normalization. The role of miR-200 in blocking cancer angiogenesis in a cancer-dependent context defines its utility as a potential therapeutic agent.

A noninvasive screening test would significantly facilitate early detection of epithelial ovarian cancer. This study used a combination of high-throughput selection and arraybased serologic detection of many antigens indicative of the presence of cancer, thereby using the immune system as a biosensor. This high-throughput selection involved biopanning of an ovarian cancer phage display library using serum immunoglobulins from an ovarian cancer patient as bait. Protein macroarrays containing 480 of these selected antigen clones revealed 65 clones that interacted with immunoglobulins in sera from 32 ovarian cancer patients but not with sera from 25 healthy women or 14 patients having other benign or malignant gynecologic diseases. Sequence analysis data of these 65 clones revealed 62 different antigens. Among the markers, we identified some known antigens, including RCAS1, signal recognition protein-19, AHNAK-related sequence, nuclear autoantogenic sperm protein, Nijmegen breakage syndrome 1 (Nibrin), ribosomal protein L4, Homo sapiens KIAA0419 gene product, eukaryotic initiation factor 5A, and casein kinase II, as well as many previously uncharacterized antigenic gene products. Using these 65 antigens on protein microarrays, we trained neural networks on two-color fluorescent detection of serum IgG binding and found an average sensitivity and specificity of 55% and 98%, respectively. In addition, the top 6 of the most specific clones resulted in an average sensitivity and specificity of 32% and 94%, respectively. This global approach to antigenic profiling, epitomics, has applications to cancer and autoimmune diseases for diagnostic and therapeutic studies. Further work with larger panels of antigens should provide a comprehensive set of markers with sufficient sensitivity and specificity suitable for clinical testing in high-risk populations. (Cancer Res 2006; 66(2): 1181-90)

Invasive micropapillary carcinoma, a tumor with highly infiltrative characteristics is defined by a distinctive cleft formation around the neoplastic cell clusters which is presumably a result of the detachment of the cells from the stroma due to as yet undetermined factors. Ultrastructural examination performed on a handful of cases demonstrated an unexpected secretory activity in the stroma-facing surface of the cells. MUC1 is a glycoprotein typically expressed in the apical surface of normal epithelial cells, responsible for maintaining lumen formation. In conventional adenocarcinomas, MUC1 expression is largely intracytoplasmic, intercellular, or apical (in glandular areas). The MUC1 expression pattern was investigated by immunohistochemical staining in invasive micropapillary carcinoma of breast (n ¼ 11), pancreas (n ¼ 5), gynecologic tract (n ¼ 11) and urinary bladder (n ¼ 10). The results were contrasted with the staining pattern in conventional carcinomas of the same organs (n ¼ 202). In all invasive micropapillary carcinoma, MUC1 expression was predominantly in the stroma-facing surface of the cell clusters (basal), accentuating the outlines of the micropapillary units by forming a distinct band on this surface. In conventional carcinoma the labeling was mostly apical in areas with lumen formation and intracytoplasmic and intercellular in the poorly differentiated areas. In conclusion, in the micropapillary pattern of invasive carcinoma, the expression of MUC1, is largely limited to the basal surface of the cells in contrast to conventional carcinomas in which MUC1 is largely apical, intracytoplasmic or intercellular. This provides support for the reversal of cell orientation as an important factor of the morphogenesis and possibly the pathogenesis of invasive micropapillary carcinoma. Since MUC1 is known to have a role in lumen formation, and has an inhibitory role in the cell to stroma interaction, it is conceivable that it is a key factor in the detachment of cells from stroma allowing for the dissection of the connective tissue and easing the spread of cells.

Background: In contrast with the decreasing incidence seen for most cancers, endometrial cancer has been increasing in the United States. We examined whether the increasing incidence and mortality from endometrial cancer are equally distributed by race/ethnicity and tumor histologic subtype.Methods: Surveillance, Epidemiology, and End Results (SEER) endometrial cancer incidence and mortality data were obtained from 2000 to 2011. Age-adjusted incidence and incidence-based mortality rates, 95% confidence intervals, and annual percent changes (APC) were calculated. Rate ratios were calculated to compare racial/ethnic groups. Five-year relative survival rates were presented to explore survival by stage at diagnosis.Results: Incidence rates for endometrial cancers are rising across all racial/ethnic groups, with the greatest APC seen among non-Hispanic black (NHB) and Asian women (APC, 2.5 for both).

A prominent feature of most cancers including Barrett’s adenocarcinoma (BAC) is genetic instability, which is associated with development and progression of disease. In this study, we investigated the role of recombinase (hsRAD51), a key component of homologous recombination (HR)/repair, in evolving genomic changes and growth of BAC cells. We show that the expression of RAD51 is elevated in BAC cell lines and tissue specimens, relative to normal cells. HR activity is also elevated and significantly correlates with RAD51 expression in BAC cells. The suppression of RAD51 expression, by short hairpin RNA (shRNA) specifically targeting this gene, significantly prevented BAC cells from acquiring genomic changes to either copy number or heterozygosity (P<0.02) in several independent experiments employing single-nucleotide polymorphism arrays. The reduction in copy-number changes, following shRNA treatment, was confirmed by Comparative Genome Hybridization analyses of the same DNA samples. Moreover, the chromosomal distributions of mutations correlated strongly with frequencies and locations of Alu interspersed repetitive elements on individual chromosomes. We conclude that the hsRAD51 protein level is systematically elevated in BAC, contributes significantly to genomic evolution during serial propagation of these cells and correlates with disease progression. Alu sequences may serve as substrates for elevated HR during cell proliferation in vitro, as they have been reported to do during the evolution of species, and thus may provide additional targets for prevention or treatment of this disease.

Although endometrial carcinoma (EC) is generally considered to have a good prognosis, over 20% of women with EC die of their disease, with a projected increase in both incidence and mortality over the next few decades. The aim of accurate prognostication is to ensure that patients receive optimal treatment and are neither overtreated nor undertreated, thereby improving patient outcomes overall. Patients with EC can be categorized into prognostic risk groups based on clinicopathologic findings. Other than tumor type and grade, groupings and recommended management algorithms may take into account age, body mass index, stage, and presence of lymphovascular space invasion. The molecular classification of EC that has emerged from the Cancer Genome Atlas (TCGA) study provides additional, potentially superior, prognostic information to traditional histologic typing and grading. This classifier does not, however, replace clinicopathologic risk assessment based on parameters other than histotype and grade. It is envisaged that molecular and clinicopathologic prognostic grouping systems will work better together than either alone. Thus, while tumor typing and grading may be superseded by a classification based on underlying genomic abnormalities, accurate assessment of other pathologic parameters will continue to be key to patient management. These include those factors related to staging, such as depth of myometrial invasion, cervical, vaginal, serosal surface, adnexal and parametrial invasion, and those independent of stage such as lymphovascular space invasion. Other prognostic parameters will also be discussed. These recommendations were developed from the International Society of Gynecological Pathologists Endometrial Carcinoma project.

Elucidation of the role of human papillomavirus (HPV) in the etiology and prognosis of squamous carcinomas of the head and neck (HNSCC) is essential to optimize prevention and treatment strategies for this disease. We analyzed 385 HNSCC tissue blocks identified through a population-based cancer registry in Metropolitan Detroit for HPV DNA using a broad-spectrum PCR technique (SPF10-LiPA25) to correlate with patient and tumor characteristics and overall survival. Overall, HPV DNA (any type) was detected in 29.4% of all HNSCC, but it was significantly more prevalent (50.6%) in oropharyngeal sites (N=81), where 90% of HPV were type 16, than in other sites. HPV prevalence (any type) in oropharyngeal sites was highest in patients with a negative smoking indicator, Caucasians, and in regional tumor stage. Likewise, only in oropharyngeal sites did patients overall positive to HPV show significantly better survival compared with HPV-negative patients, notably among those who had been irradiated. The best and the worst survival from cancer in oropharyngeal sites were found, respectively, among HPV-positive patients with negative smoking indicator and among HPV-negative patients with positive smoking indicator. The results of the present study revealed that the presence of HPV DNA was associated with patients’ specific characteristics and better overall survival exclusively in oropharyngeal sites. To define the fraction of HNSCC preventable by HPV vaccination or amenable to less aggressive treatment, however, tobacco exposure and HPV markers other than DNA presence need to be taken into account.